The kinetic analysis of ligand receptor interactions suggests that receptors and G proteins exist in rapidly coupling and slowly coupling pools in membrane surfaces. To determine whether receptors in the slowly coupling pools were physically precoupled to G proteins or coupled rapidly, we have examined a number of peptides for their ability to interfere with receptor-G protein coupling in broken cell preparations. These peptides represent portions of receptors and G proteins in regions alledged to mediate coupling during signal transduction. Although these peptides have been shown to affect the interactions between the purified proteins, they do not seem to affect the interactions of the receptors and G protein in the cell membranes. This observation leads us to suggest that the receptors and G proteins are physically precoupled, supporting the model derived from kinetic analysis. The next phase of the project will involve a description of the underlying biochemistry of signal transduction. The kinetic methods will combine rapid mix flow cytometry with biochemisty to evaluate the evolution of receptors among different macromolecular complexes. These receptor complexes will be isolated by immunopurification of epitope tagged receptor and the transduction molecules in the comples will be identified. We will attempt to develop a mathematial model from the kinetic methods to account for the biochemical steps in receptor activation and desensitization.